Clinical Significance And Molecular Pathogenesis Of Hepa

B型肝炎病毒（HBV）感染的结局是多种病毒与宿主相互作用的复杂结果。虽然宿主反应可能起主要作用，但这一过程通常取决于各种病毒适应机制。通常，病毒基因组关键区域的病毒突变是这些适应机制的结果。我们的实验室已经确定并表征了与某些HBV毒株的变异生物学行为相关的特定病毒突变。在与暴发性肝炎相关的HBV毒株中鉴定了HBV核心启动子中的两个突变，由于前基因组RNA进入核心颗粒的病毒掺入增加，导致复制高度增强。我们最近的出版物表明，自然发生的突变，影响一种新的遗传元件可能会影响病毒的共转录或转录后机制，导致增强核心合成和病毒复制。作为该项目的第二个方面，我们已经开始研究HBV感染中毛玻璃样肝细胞的发病机制。磨玻璃样肝细胞是慢性B型肝炎病毒（HBV）感染的独特组织学特征。这些肝细胞被抗-HBs和抗-preS 1强染色。电镜研究表明，大量的HBV包膜蛋白聚集在扩张的囊泡内，推测这些囊泡来自内质网。在转基因小鼠中，大表面蛋白的过度表达导致毛玻璃样肝细胞。大表面蛋白的pre-S1区域已被证明调节HBsAg的组装、加工和分泌。因此，我们假设前S1的突变形式影响这一正常的分泌途径，并负责毛玻璃肝细胞。为了研究这种可能性，我们检测了HBV感染患者的HBV前S1区序列，肝活检时有明显的毛玻璃样肝细胞。为了分析单个毛玻璃样肝细胞的病毒群体，我们使用激光捕获显微切割技术从活检标本中分离单个肝细胞。分别收获抗-HBs染色阳性的毛玻璃样肝细胞以及外观正常的肝细胞，并对其HBV DNA进行序列分析。对一名患者的初步分析显示，大多数来自毛玻璃肝细胞的病毒分离株含有独特的前S1突变序列。相比之下，对照肝细胞仅具有WT序列。这个前S突变体的功能研究表明，不寻常的电泳特性和非典型的细胞分布，可能有生物学意义的毛玻璃细胞的形成。正在大量患者中进行其他研究，以分析这些前S1突变的功能效应。明确与自然发生的HBV突变体感染相关的肝病变异表现的分子基础可能有助于进一步了解HBV感染的发病机制。

The outcome of hepatitis B virus (HBV) infection results from complicated interplay among a variety of virus-host interactions. Although host responses likely play a major role, this process often depends on a variety of viral adaptive mechanisms. Frequently, viral mutations in critical regions of viral genome are the results of these adaptive mechanisms. Our laboratory has identified and characterized specific viral mutations associated with variant biological behaviors of certain HBV strains. Two mutations in the HBV core promotor were identified in a HBV strain associated with fulminant hepatitis leading to highly enhanced replication as a result of increased viral encapsidation of pregenomic RNA into the core particles. Our recent publications suggest that naturally occurring mutations affecting a novel genetic element may influence viral encapsidation by a co- or post-transcriptional mechanism resulting in enhanced core synthesis and viral replication. As a second aspect of this project, we have initiated studies into the pathogenesis of ground glass hepatocytes in HBV infection. Ground glass hepatocyte is an unique histological feature of chronic hepatitis B virus (HBV) infection. These hepatocytes are stained strongly with anti-HBs and anti-preS1. EM studies show that large amounts of HBV envelope proteins accumulate within dilated vesicles presumably derived from the endoplasmic reticulum. In transgenic mice, overexpression of large surface protein leads to ground glass hepatocytes. The pre-S1 region of the large surface protein has been shown to regulate assembly, processing and secretion of HBsAg. We therefore hypothesize that a mutant form of pre-S1 affects this normal secretory pathway and is responsible for ground glass hepatocytes. To investigate this possibility, we examined HBV sequences spanning the pre-S1 region from HBV infected patients with evident ground glass hepatocytes on liver biopsy. To analyze the viral population of single ground glass hepatocytes, we used the technique of laser capture microdissection to isolate individual hepatocytes from the biopsy specimen. Ground glass hepatocytes that stained positively with anti-HBs as well as normal appearing hepatocytes were harvested individually and their HBV DNA subjected to sequence analysis. Preliminary analysis of one patient revealed that the majority of viral isolates from the ground glass hepatocytes contained a unique pre-S1 mutant sequence. In contrast, the control hepatocytes had exclusively WT sequence. Functional study of this pre-S mutant demonstrated unusual electrophoretic properties and atypical cellular distribution that may have biological implications in the formation of ground glass cells. Additional studies are being pursued in a large number of patients and to analyze the functional effect of these pre-S1 mutations. Defining the molecular basis of variant manifestations of liver disease associated with infection by naturally occurring HBV mutants may contribute to further understanding of the pathogenesis of HBV infection.